红鲫胚胎发育过程中Sox8基因对壬基酚胁迫的响应

Response of Sox8 gene to nonylphenol stress during red crucian carp(Carassius auratus red var.) development

  • 摘要: 【目的】明确红鲫SRY相关高迁移率族盒蛋白8基因(Sox8)表达模式及壬基酚(NP)暴露对红鲫胚胎发育过程中Sox8基因表达的影响,为揭示NP胁迫下红鲫胚胎发育畸形的分子机制提供科学依据。【方法】运用RACE克隆红鲫Sox8基因cDNA全长序列,通过ProtParam、ProtScale、InterPro、PredictProtein、DeepTMHMM等在线软件进行生物信息学分析,利用实时荧光定量PCR检测红鲫Sox8基因在各成体组织及不同胚胎发育阶段的表达情况,以及不同浓度NP暴露对红鲫胚胎期Sox8基因表达的影响。【结果】红鲫Sox8基因c DNA序列全长2163 bp,其开放阅读框(ORF)1176bp,共编码391个氨基酸残基;红鲫SOX8蛋白相对分子量为43798.30Da,理论等电点(pI)为6.90,脂溶指数为49.92,不稳定指数为66.04,总平均亲水性指数(GRAVY)为-1.026,其N端含有Sox_N二聚化结构域和HMG-box结构域;基于SOX8氨基酸序列相似性构建的系统发育进化树显示红鲫与鲫的亲缘关系最近。红鲫Sox8基因在各成体组织中广泛表达,以脑组织中的Sox8基因相对表达量最高,显著高于其他组织(P<0.05,下同);在红鲫胚胎发育过程中,Sox8基因的表达从受精后到14体节期呈逐渐下降趋势,但从21体节期开始Sox8基因相对表达量显著增高,直到孵化期始终维持在较高水平。在大多数情况下,NP暴露能显著影响红鲫胚胎发育过程中Sox8基因的表达,特别是14体节期后的发育阶段,NP暴露主要是不同程度地激活Sox8基因表达。【结论】Sox8基因表达在红鲫成体中具有组织特异性,且在胚胎发育过程中呈先降低后升高的变化趋势。NP暴露能显著影响红鲫胚胎发育过程中Sox8基因的表达,14体节期后Sox8基因表达水平升高可能是NP致畸红鲫胚胎的重要原因之一。

     

    Abstract: 【Objective】To elucidate the expression pattern of the SRY-related high mobility group box protein 8 gene(Sox8) in red crucian carp(Carassius auratus red var.) and the impact of nonylphenol(NP) exposure on Sox8 gene expression during embryonic development, providing a scientific basis for uncovering the molecular mechanisms of NPinduced embryonic developmental deformities in red crucian carp. 【Method】The full-length cDNA sequence of Sox8 gene from red crucian carp was cloned using RACE technique. Bioinformatics analysis were conducted using online tools such as ProtParam, ProtScale, InterPro, PredictProtein and DeepTMHMM. Real-time fluorescence quantitative PCR was employed to detect the expression of Sox8 gene in various adult tissues and at different embryonic developmental stages, as well as the effect of different NP concentrations exposure on Sox8 gene expression during the embryonic stage of red crucian carp. 【Result】The full-length cDNA sequence of Sox8 gene in red crucian carp was 2163 bp, with an open reading frame(ORF) of 1176 bp encoding 391 amino acid residues. The relative molecular weight of the SOX8 protein was 43798.30 Da, with a theoretical isoelectric point(pI) of 6.90, a lipophilicity index of 49.92, an instability index of 66.04, and a grand average of hydropathicity(GRAVY) of-1.026. The N-terminus contained a Sox_N dimerization domain and an HMG-box domain. Phylogenetic analysis based on the SOX8 amino acid sequences revealed a close relationship between C. auratus red var. and Carassius carassius(common crucian carp). The Sox8 gene was broadly expressed in various adult tissues of red crucian carp, with the highest relative expression in the brain, which was significantly higher than that in other tissues(P<0.05, the same below). During embryonic development, Sox8 gene expression decreased gradually from post-fertilization to the 14-somite stage, but significantly increased from the 21-somite stage, maintained at a high level until the hatching stage. Under most conditions, NP exposure significantly affected Sox8 gene expression during embryonic development of red crucian carp, particularly activating Sox8 gene expression to varying degrees after the 14-somite stage. 【Conclusion】Sox8 gene expression exhibits tissue specificity in adult red crucian carp and presents a trend of initial decrease followed by increase during embryonic development. NP exposure significantly affects Sox8 gene expression during embryonic development, with the increased expression of Sox8 gene after the 14-somite stage potentially contributing to NP-induced embryonic deformities in red crucian carp.

     

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